Mycobacterium avium causes infection in both immunocompromised patients and in healthy individuals. The chiefly encountered infections are in the respiratory tract, and disseminated. The ability of the bacterium to survive in different environments in the host requires tight gene regulation. During the past 4 years of this grant, we have unveiled a number of the complex aspects of M. avium interaction with the host, from the interaction between the bacterium and the host mucosa, to the ability to survive in macrophages and to disseminate. Several virulence genes were identified and their functions characterized. Many of the genes allow the bacterium to enter intestinal epithelial cells, and others are required for survival within macrophages. The long-term goal of this project is to understand the mechanisms of interaction between the pathogen and host cells. Our submission now contains proposed studies to explore the relationship between M. avium and macrophages. Our central hypothesis is that M. avium has specific strategies to subvert the host cells and that becoming aware of them will allow for improved means of prevention and treatment of M. avium-related diseases. We propose to continue this work during the next funding period by evaluating: 1. M. avium interference with macrophage killing mechanisms: (a) studying bacterial genes that participate in modifying/interfering with the mycobacterial vacuole formation upon uptake into macrophages; (b) studying the mechanisms of action of genes associated with bacterial resistance to macrophage killing mechanisms; 2. The mechanisms by which M. avium leaves apoptotic macrophages and spreads to surrounding macrophages.